Transfection of NRVMs with RhoA antisense oligonucleotide attenuated stretch-induced FAK and ERK1/2 phosphorylation and expression of -myosin heavy chain mRNA. Similar results were seen in cells transfected with FAK antisense oligonucleotide. These findings demonstrate that RhoA/ROCK signaling plays a crucial role in stretch-induced FAK phosphorylation, presumably by coordinating upstream events operationally linked to the actin cytoskeleton. mechanical stress; hypertrophy; cell signaling INCREASED BIOMECHANICAL STRESS can drive changes in cardiac myocytes that are implicated in myocardial hypertrophy and failure (7,19). Numerous signal transduction pathways have been shown to be activated in cardiac myocytes subjected to mechanical stimuli (35). Signals originating from multiple pathways converge intracellularly, leading to altered gene expression and protein synthesis, which result in the hypertrophic growth of cardiac myocytes. However, the mechanism by which mechanical forces are sensed and converted to biochemical signals remains largely unknown. Recent developments in this field indicate that the integrity of structures such as the Z disk, costamere, and intercalated disk is critically important to the ability of cardiac cells to appropriately respond to mechanical stress (4,11,21,32,37,41). It has been hypothesized that such structures participate in monitoring of mechanical force and in communication of strain to signaling molecules in cardiac myocytes (12,29,37).Focal adhesion kinase (FAK), a tyrosine kinase linked to integrin signaling (15,24,42), has been shown to be rapidly activated by mechanical stimuli in cardiac myocytes (2,5,9,12,13,23,34,39). Several lines of evidence support a role for FAK in the regulation of early gene transcription in response to hypertrophic agonists and mechanical stress (10,22,28,38,39), indicating that this kinase may coordinate the convergence of multiple signaling pathways involved in the hypertrophic growth of cardiac myocytes. However, the molecular mechanism responsible for FAK activation by mechanical stress in cardiac myocytes remains elusive. We recently showed (12, 39) that FAK activation by mechanical stress is accompanied by its aggregation at myofilaments, Z disks, and costameres, implying that this kinase might be directly activated by mechanical stress in cardiac myocytes. On the other hand, FAK activation in neonatal rat ventricular myocytes (NRVMs) by agonists such as endothelin has been demonstrated (16) to be dependent on activation of the RhoA/Rho-associated coiled coil-containing protein kinase (ROCK) signaling pathway, which drives the assembly and rearrangement of actin filaments. The demonstration that cytochalasin D, a potent inhibitor of actin polymerization, markedly attenuated endothelininduced FAK phosphorylation (16) revealed the importance of the assembly of actin filaments in FAK activation by this agonist. Similarly, FAK activation by mechanical stress has been suggested to depend on a cooperative interaction with actin filaments (13,39,...